Visible Floral Bud Research Articles

Virus-induced gene silencing (VIGS) for functional analysis of genes involved in the regulation of anthocyanin biosynthesis in the perianth of Phalaenopsis-type Dendrobium hybrids

• We firstly conducted Cymbidium mosaic virus induced gene silencing in Phalaenopsis -type Dendrobium hybrids (Den-Phals). • The regulatory gene DhMYB2 for anthocyanin biosynthesis was considered as a visible marker in VIGS study for floral genes function in Den-Phals. • DhMYB2 was demonstrated as the key regulatory gene for the anthocyanin accumulation in sepals, petals and mid-lobes of labella in flowers of Den-Phals. Virus-induced gene silencing (VIGS) is an efficient way to study floral gene functions of orchids that have long life cycle and inefficient genetic transformation. However, reports of the use of the VIGS technique for Dendrobium spp. are scarce. To explore the applicability of Cymbidium mosaic virus (CymMV)-based VIGS in Phalaenopsis -type Dendrobium hybrids (Den-Phals), the regulator gene DhMYB2 of anthocyanin biosynthesis was used as a target gene. Several variables including the length of DNA insertion, the location of the cDNA, and the developmental state of inflorescences were examined. The results suggested that a CymMV-based VIGS system could be used with Den-Phals to silence DhMYB2 up to 98%, resulting in the loss of anthocyanin coloration in sepals, petals and mid-lobes of labella and down regulation of the expression levels of major genes in the anthocyanin biosynthetic pathway (ABP). The sizes of the DNA insert for silencing were between 120 and 200 bp. The most obvious phenotype was obtained when using the 5′ terminus of the coding region of DhMYB2 cDNA. The plants of a younger development stage of inflorescence with no more than four visible floral buds were suitable for Agro-infiltration. The present study demonstrated the feasibility of this CymMV-based VIGS system using DhMYB2 as the visible marker gene for the functional analysis of floral genes and illuminated the regulatory effects of DhMYB2 on the floral pigmentation in Den-Phals.

Ectopic expression of an Eriobotrya japonica APETALA3 ortholog rescues the petal and stamen identities in Arabidopsis ap3-3 mutant

Apetala3(AP3) encodes a floral homeotic class B-function MADS-box protein and plays crucial roles in petal and stamen development. To better understand the functional roles of AP3 orthologs in Eriobotrya, we isolated and identified an AP3 ortholog, referred to as EjAP3, from Eriobotrya japonica. Analyses of protein sequence and phylogenetic tree showed that the EjAP3 was assigned to the rosids euAP3 lineage and included a distinctive PI-derived and euAP3 motifs at the C-terminal domain. Subcellular localization of EjAP3 was determined to be in the nucleus. Expression analysis suggested that EjAP3 expression was restricted only in petals and stamens, but not in sepals and carpels. Importantly, during the floral development, EjAP3 expression level was the highest at the stage of visible floral bud. Furthermore, ectopic expression of EjAP3 in Arabidopsis ap3-3 mutant rescued the second whorl petals and the third whorl stamens. The expression pattern and function characterization of EjAP3 contribute to better understand the roles of AP3 orthologs in Eriobotrya.

CO2 Enrichment with Higher Light Level Improves Flowering Quality of Phalaenopsis Queen Beer ‘Mantefon’

We investigated the flowering and photosynthetic responses of Phalaenopsis Queen Beer ‘Mantefon’ to CO2 enrichment and light level. Potted plants at the flowering stage were supplied with low light level (LL; 90 ± 10 μmol m−2 s−1) or high light level (HL; 260 ± 40 μmol m−2 s−1) using three−wave cool white fluorescent lamps and metal halide lamp and different levels of CO2. Carbon dioxide was supplied to the plants, which perform crassulacean acid metabolism, during their dark period at three concentrations: 400 (control), 800, and 1200 μmol mol−1 CO2. More flowers and lateral branches were produced by plants grown under HL than the control plants grown under LL, and there were more visible floral buds in the plants grown under HL than those grown LL, regardless of CO2 concentration. Net CO2 assimilation rate was higher in plants exposed to 800 and 1200 μmol mol−1 CO2 under LL and 1200 μmol mol−1 CO2 than the control plants under LL and HL. We conclude that photosynthesis is enhanced in CO2-enriched plants, and lateral branch and floral bud production are maximized at the high light level used here in Phalaenopsis Queen Beer ‘Mantefon’.

Adaptive responses of quinoa to diverse agro-ecological environments along an altitudinal gradient in North West Argentina

Quinoa is an important Andean grain crop grown in a wide range of tropical and temperate environments. Time to flowering is an important trait determining grain yield. This work aimed to understand how responses to photoperiod and temperature might alter plant leaf and floral development. To assess the likely degree of G×E interactions, eleven quinoa accessions from a wide range of environments of origin within Northwest Argentina were grown in several sowing dates over two seasons at a high altitude site. In a third season at a low altitude site, a subset of six accessions planted in pots in the field was exposed to two artificially extended and a control (natural) photoperiod. Time to the appearance of floral buds and anthesis were recorded as was leaf number. A photothermal model developed for quinoa was used to compare responses to photoperiod. Plant development rates to visible floral buds and anthesis stages and phyllochron varied widely amongst accessions and across environments within a short day response though phyllochron varied mostly during the reproductive phase. There was a very strong association between time to flowering and altitude of origin (r=−0.98), mean temperature of the wettest quarter (r=0.98) and Normalized Difference Vegetation Index values (r=0.73). Photoperiod sensitivity was higher for accessions from the lowlands (normally late flowering), while temperature sensitivity was greatest for accessions from the highlands (early flowering); most variation for these traits detected at the species level was found in North West Argentina. Genotype by environment interactions for yield were related to the traits examined in this study and considering their high heritability it is suggested that quinoa breeding programs targeted for specific adaptation to a wide range of environments can be developed from this germplasm.

Multispectral detection of floral buds for automated thinning of pear

Thinning of pome and stone fruit involves the reduction of tree crop load in order to regulate fruit set and quality. As it is typically carried out through manual labor, thinning comprises a large part of a grower’s production costs. Mechanized thinning has been shown to be a cost-effective alternative but the performance of existing thinning devices needs to be further improved by taking the variation in bearing capacity of the individual trees into account.In this work, a multispectral camera system is developed to detect the floral buds of pear (cv. Conference) during the growth stages prior to bloom. During a two-year field trial, the multispectral system was used to measure orchard scenes in six distinct optical wavebands under controlled illumination. These wavebands are situated in the visible and near infrared region of the spectrum and were selected based on hyperspectral laboratory measurements described in previous work.The recorded multispectral images were converted to a database containing the spatial–spectral signatures of the objects present in the orchard. Subsequently, canonical correlation analysis was applied to create a spectral discriminant model that detects pixels originating from floral buds. This model was then applied to the recorded data after which an image analysis algorithm was designed and optimized to predict the number of floral buds. In total, approximately 87% of the visible floral buds were detected correctly with a low false discovery rate (<16%). Therefore, it is expected that the multispectral sensor can be used to improve the efficiency of existing thinning devices. Additionally, it could as well be used as a stand-alone sensor for early-season yield estimation.

Photoperiodic control of FT-like gene ClFT initiates flowering in Chrysanthemum lavandulifolium

The FLOWERING LOCUS T (FT) gene plays crucial roles in regulating the transition from the vegetative phase to the reproductive phase. In this study, we isolated an FT homologous gene (denoted as ClFT) from Chrysanthemum lavandulifolium. The sequencing analysis indicated that the promoter of the ClFT gene contains many elements, such as light response, abscisic acid, drought-inducibility response and CIRCADIAN clock elements. The expression patterns of ClFT in different tissues/organs at different developmental stages and its responses to different photoperiods were observed. ClFT is expressed in all tested organs/tissues, with the highest expression level being observed in the leaves of plants with visible floral buds under the short day (SD) condition. Next, we studied the rhythmic expression of ClFT during different photoperiod treatments and found that the level of ClFT increases with additional hours of continuous dark. ClFT accumulates when the continuous dark period is 12 h, regardless of the duration of light period. The ectopic expression of the ClFT gene in wild type Arabidopsis (Col-0) results in early flowering, with high expression levels of endogenous LFY and SOC1 being observed in transgenic Arabidopsis. All results indicated that the ClFT gene plays an evolutionarily conserved role in promoting flowering in inductive short days in C. lavandulifolium and that this gene could serve as a vital target for the genetic manipulation of flowering time in chrysanthemums.

Modelling temperature, photoperiod and vernalization responses of Brunonia australis (Goodeniaceae) and Calandrinia sp. (Portulacaceae) to predict flowering time

Crop models for herbaceous ornamental species typically include functions for temperature and photoperiod responses, but very few incorporate vernalization, which is a requirement of many traditional crops. This study investigated the development of floriculture crop models, which describe temperature responses, plus photoperiod or vernalization requirements, using Australian native ephemerals Brunonia australis and Calandrinia sp. A novel approach involved the use of a field crop modelling tool, DEVEL2. This optimization program estimates the parameters of selected functions within the development rate models using an iterative process that minimizes sum of squares residual between estimated and observed days for the phenological event. Parameter profiling and jack-knifing are included in DEVEL2 to remove bias from parameter estimates and introduce rigour into the parameter selection process. Development rate of B. australis from planting to first visible floral bud (VFB) was predicted using a multiplicative approach with a curvilinear function to describe temperature responses and a broken linear function to explain photoperiod responses. A similar model was used to describe the development rate of Calandrinia sp., except the photoperiod function was replaced with an exponential vernalization function, which explained a facultative cold requirement and included a coefficient for determining the vernalization ceiling temperature. Temperature was the main environmental factor influencing development rate for VFB to anthesis of both species and was predicted using a linear model. The phenology models for B. australis and Calandrinia sp. described development rate from planting to VFB and from VFB to anthesis in response to temperature and photoperiod or vernalization and may assist modelling efforts of other herbaceous ornamental plants. In addition to crop management, the vernalization function could be used to identify plant communities most at risk from predicted increases in temperature due to global warming.

Juvenility and flowering of Brunonia australis (Goodeniaceae) and Calandrinia sp. (Portulacaceae) in relation to vernalization and daylength

The time at which plants are transferred to floral inductive conditions affects the onset of flowering and plant morphology, due to juvenility. Plants of Brunonia australis and Calandrinia sp. were used to investigate whether Australian native ephemeral species show a distinct juvenile phase that can be extended to increase vegetative growth and flowering. The juvenile phase was quantified by transferring seedlings from less inductive (short day and 30/20°C) to inductive (vernalization or long day) conditions at six different plant ages ranging from 4 to 35 d after seed germination. An increase in days to first visible floral bud and leaf number were used to signify the end of juvenility. Brunonia australis was receptive to floral inductive long day conditions about 18-22 d after seed germination, whereas plants aged 4-35 d appeared vernalization sensitive. Overall, transferring plants of B. australis from short to long day conditions reduced the time to anthesis compared with vernalization or constant short day conditions. Calandrinia sp. showed a facultative requirement for vernalization and an insensitive phase was not detected. Floral bud and branch production increased favourably as plant age at time of transfer to inductive conditions increased. Younger plants showed the shortest crop production time. Both species can perceive the vernalization floral stimulus from a very young age, whereas the photoperiodic stimulus is perceived by B. australis after a period of vegetative growth. However, extending the juvenile phase can promote foliage development and enhance flower production of both species.

Vernalization promotes flowering of a heat tolerant Calandrinia while long days replace vernalization for early flowering of Brunonia

The flowering responses of Brunonia australis (blue pincushion) and Calandrinia sp. to vernalization, photoperiod, temperature and plant age were investigated to provide a foundation for manipulating flowering in these potential potted plants. Plants were vernalized at 4.8 °C for 0, 3 or 6 weeks at the plant age of 1–4 or 8–14 leaves. Following vernalization, plants were grown at 25/10 or 35/20 °C (day/night) under short days (11 h, ambient daylight averaged 380 ± 44 μmol m −2 s −1) or long days (16 h) provided by an additional 5 h night break (21:00–2:00 h at <4.5 μmol m −2 s −1 from incandescent lamps), for 85 days. This is the first work to investigate flowering of these ornamental species. Both species showed enhanced flowering following vernalization and a quantitative requirement for long days. The reduction of the time until the first visible inflorescence ( Brunonia) or flower ( Calandrinia) buds by 8–13 days was affected by vernalization for 3 or 6 weeks, respectively. Long days were effective for reducing the time to first visible floral bud and increasing the number of inflorescence or flowers per plant for both species. For Brunonia, LDs replaced vernalization when applied to plants with 1–4 leaves. Raising temperature from 25/10 to 35/20 °C increased the number of flowers per plant of Calandrinia by 2–2.5-fold for plants with 1–4 or 8–14 leaves respectively.

Development and Abortion of Flowers in Capsicum annuum Exposed to High Temperatures

Reduction of floral number in Capsicum annuum has been observed during growth at high temperature. To determine whether decreased flower production or increased flower abscission is a direct response to high temperatures or a response to water stress induced by high temperatures, we compared flowers and fruit produced and flowers aborted to leaf growth rate, osmotic potential, stomatal conductance, and chlorophyll fluorescence of two cultivars. To determine the stage(s) of floral development that are most sensitive to high temperatures, flower buds were wax-embedded and examined at each stage of development during heat treatment. Rate of floral development also was examined. At first visible floral bud initiation, plants were transferred to each of three controlled environment growth chambers with set temperatures and vapor pressure deficits (VPD) of 25°C, 1.1 kPa; 33°C, 1.1 kPa; and 33°C, 2.1 kPa. Flower bud production and leaf growth rate were not significantly affected by high temperatures. Pepper fruit set, however, was inhibited at 33°C at either VPD. Preliminary water relations data suggested that water potentials were more negative under high temperature conditions. Differences in leaf fluorescence were statistically significant for temperature treatments, but not for VPD. Temperature is the primary factor in the decrease of fruit production in pepper. Decreased production is due to flower abortion and not to decreased flower initiation or plant growth.

2502 The pincushion grid illusion: a kind of the neon spreading illusion?

The flowering responses of Brunonia australis (blue pincushion) and Calandrinia sp. to vernalization, photoperiod, temperature and plant age were investigated to provide a foundation for manipulating flowering in these potential potted plants. Plants were vernalized at 4.8 °C for 0, 3 or 6 weeks at the plant age of 1–4 or 8–14 leaves. Following vernalization, plants were grown at 25/10 or 35/20 °C (day/night) under short days (11 h, ambient daylight averaged 380 ± 44 μmol m−2 s−1) or long days (16 h) provided by an additional 5 h night break (21:00–2:00 h at <4.5 μmol m−2 s−1 from incandescent lamps), for 85 days. This is the first work to investigate flowering of these ornamental species. Both species showed enhanced flowering following vernalization and a quantitative requirement for long days. The reduction of the time until the first visible inflorescence (Brunonia) or flower (Calandrinia) buds by 8–13 days was affected by vernalization for 3 or 6 weeks, respectively. Long days were effective for reducing the time to first visible floral bud and increasing the number of inflorescence or flowers per plant for both species. For Brunonia, LDs replaced vernalization when applied to plants with 1–4 leaves. Raising temperature from 25/10 to 35/20 °C increased the number of flowers per plant of Calandrinia by 2–2.5-fold for plants with 1–4 or 8–14 leaves respectively.

The pincushion grid illusion: a kind of the neon spreading illusion?

The flowering responses of Brunonia australis (blue pincushion) and Calandrinia sp. to vernalization, photoperiod, temperature and plant age were investigated to provide a foundation for manipulating flowering in these potential potted plants. Plants were vernalized at 4.8 °C for 0, 3 or 6 weeks at the plant age of 1–4 or 8–14 leaves. Following vernalization, plants were grown at 25/10 or 35/20 °C (day/night) under short days (11 h, ambient daylight averaged 380 ± 44 μmol m−2 s−1) or long days (16 h) provided by an additional 5 h night break (21:00–2:00 h at <4.5 μmol m−2 s−1 from incandescent lamps), for 85 days. This is the first work to investigate flowering of these ornamental species. Both species showed enhanced flowering following vernalization and a quantitative requirement for long days. The reduction of the time until the first visible inflorescence (Brunonia) or flower (Calandrinia) buds by 8–13 days was affected by vernalization for 3 or 6 weeks, respectively. Long days were effective for reducing the time to first visible floral bud and increasing the number of inflorescence or flowers per plant for both species. For Brunonia, LDs replaced vernalization when applied to plants with 1–4 leaves. Raising temperature from 25/10 to 35/20 °C increased the number of flowers per plant of Calandrinia by 2–2.5-fold for plants with 1–4 or 8–14 leaves respectively.

2501 Representation of similarity among 3D shapes in the monkey visual system

The flowering responses of Brunonia australis (blue pincushion) and Calandrinia sp. to vernalization, photoperiod, temperature and plant age were investigated to provide a foundation for manipulating flowering in these potential potted plants. Plants were vernalized at 4.8 °C for 0, 3 or 6 weeks at the plant age of 1–4 or 8–14 leaves. Following vernalization, plants were grown at 25/10 or 35/20 °C (day/night) under short days (11 h, ambient daylight averaged 380 ± 44 μmol m−2 s−1) or long days (16 h) provided by an additional 5 h night break (21:00–2:00 h at <4.5 μmol m−2 s−1 from incandescent lamps), for 85 days. This is the first work to investigate flowering of these ornamental species. Both species showed enhanced flowering following vernalization and a quantitative requirement for long days. The reduction of the time until the first visible inflorescence (Brunonia) or flower (Calandrinia) buds by 8–13 days was affected by vernalization for 3 or 6 weeks, respectively. Long days were effective for reducing the time to first visible floral bud and increasing the number of inflorescence or flowers per plant for both species. For Brunonia, LDs replaced vernalization when applied to plants with 1–4 leaves. Raising temperature from 25/10 to 35/20 °C increased the number of flowers per plant of Calandrinia by 2–2.5-fold for plants with 1–4 or 8–14 leaves respectively.

Development rate of young greenhouse rose plants ( Rosa hybrida) rooted from cuttings in relation to temperature and irradiance

The development rate of 13 genotypes of greenhouse roses was observed for young plants rooted from cuttings and grown in a phytotron or greenhouse. The development was divided into three stages from pinching to anthesis: Stage 1, shoot 0 cm (pinching) to 1 cm long (bud break) (0 → 1); Stage 2, shoot 1 cm long to visible floral bud (B) (1 → B); Stage 3, visible floral bud to anthesis ( B → F). In the phytotron, with photosynthetically active radiation (PAR) from 0.65 to 2.60 MJ m −2 day −1, each phase of Rosa hybrida cultivar ‘Sonia’ had different responses to irradiance and mean air temperature ( T). The duration ( d) of Stage 1 was primarily a function of T, that of Stage 2 was a power function of PAR and a parabolic or hyperbolic function of T: d 1 → B = k × PAR x × f( T). The duration of Stage 3 was much less sensitive to PAR, and was a parabolic or hyperbolic function of T. In the greenhouse, with miniplants grown at different seasons, these equations for the effects of T and PAR were applied and validated with at least the ten varieties which rooted normally during the cutting process, and which flowered normally even in winter. Owing to a strong interaction of PAR and T in the greenhouse, the varietal PAR effects should be determined principally in a phytotron, then the T effect may be determined in a series of greenhouse experiments.

Genotypic Variation and Classification of Cowpea for Reproductive Responses to High Temperature under Long Photoperiods

Hot weather can substantially reduce pod production by cowpea [Vigna unguiculata (L.) Walp.]. The objectives were to characterize and classify cowpea based on their responses to heat during reproductive development. Diverse cowpea accessions were grown over six summers in Imperial Valley, CA under hot temperatures (mean daily max/min 41/24 °C) and long days (14 h, 51 min decreasing to 12 h, 18 min). Selected genotypes were grown at Riverside, CA, under more optimal temperatures (35/17 °C) and the same daylengths. At Imperial Valley, genotypic variation was observed for duration to first macroscopic floral bud and extent of floral bud abortion, peduncle elongation, flower production, and podding. At Riverside, similar genotypic variation occurred for days to first floral bud, but subsequent reproductive development was normal. Controlled environment studies in growth chambers showed that the detrimental effects induced in sensitive genotypes by high temperature were enhanced by long photoperiods. Based on these differing reproductive responses, the accessions were classified into eight groups. Heat‐tolerant accessions in Group I exhibited normal peduncle elongation, early flowering, and produced many flowers and pods. Accessions in Groups II and III performed similarly but set few pods (Group II) or no pods (Group III). Accessions in Groups IV through VII differed in duration to first floral bud, with Group VII being the latest and exhibiting substantial bud abortion, suppressed peduncle elongation, and no flowers. Group VIII accessions produced no visible floral buds. Cowpea developed in different regions and for specific seasons within a region fell into discrete classification groups, indicating that the system has a genetic and ecological basis. This classification system is useful for characterizing the types of heat responses present in cowpea genotypes and for selecting appropriate combinations of parents for breeding heat‐tolerant cultivars for different production zones and seasons.

Abortion of Floral Buds in Acomastylis rossii Plants Exposed to Artificial Acid Mists in Alpine Tundra

Plants of Acomastylis rossii were treated over a three-year period with artificial acid mists of pH 2.5, 3.5, or 4.5 prepared with sulfuric acid or nitric acid. Treatments were made in the field twice weekly for eight weeks during each of the three growing seasons. Significant decreases in the percentage of plants flowering were noted in each year of the study in plants treated with sulfuric acid mist at pH 2.5. Significant decreases in flowering with sulfuric acid at pH 3.5 and in leaf number with sulfuric acid at pH 2.5 and 3.5 occurred in the third year of treatments as well. Some individual plants flowered in one, two, or three of the years, indicating that new floral primordia were being produced during the treatment period. Plants that flowered produced viable, germinable seeds. No effects of nitric acid mists were noted during the study period. Treatments with citrate buffer solutions (pH 6.2) used to determine if the plants were responding to sulfate independently of pH showed no significant differences for any of the measurements taken. The observed decreases in flowering are apparently a sulfuric acid effect and are not attributable to hydrogen ions or sulfate ions independently of each other. Experiments with plants that had visible floral buds indicate that plants aborted the floral structures in response to direct contact of buds with solution from the acid mists. The growth form of the plants enhances contact by causing pooling of the solutions on top of the developing buds in pockets formed by the basal leaves.

ABORTION OF FLORAL BUDS IN ACOMASTYLIS ROSSII PLANTS EXPOSED TO ARTIFICIAL ACID MISTS IN ALPINE TUNDRA

Plants of Acomastylis rossii were treated over a three‐year period with artificial acid mists of pH 2.5, 3.5, or 4.5 prepared with sulfuric acid or nitric acid. Treatments were made in the field twice weekly for eight weeks during each of the three growing seasons. Significant decreases in the percentage of plants flowering were noted in each year of the study in plants treated with sulfuric acid mist at pH 2.5. Significant decreases in flowering with sulfuric acid at pH 3.5 and in leaf number with sulfuric acid at pH 2.5 and 3.5 occurred in the third year of treatments as well. Some individual plants flowered in one, two, or three of the years, indicating that new floral primordia were being produced during the treatment period. Plants that flowered produced viable, germinable seeds. No effects of nitric acid mists were noted during the study period. Treatments with citrate buffer solutions (pH 6.2) used to determine if the plants were responding to sulfate independently of pH showed no significant differences for any of the measurements taken. The observed decreases in flowering are apparently a sulfuric acid effect and are not attributable to hydrogen ions or sulfate ions independently of each other. Experiments with plants that had visible floral buds indicate that plants aborted the floral structures in response to direct contact of buds with solution from the acid mists. The growth form of the plants enhances contact by causing pooling of the solutions on top of the developing buds in pockets formed by the basal leaves.

Phenology and reproductive physiology of Thalassia testudinium from the Western tropical Atlantic

Phenological investigations in natural seagrass beds of Texas, Florida and St. Croix, U.S. Virgin Islands, and reproductive physiology studies under controlled conditions suggest that flowering in Thalassia testudinum Banks ex König is related to the temperature progressions that follow winter minima. Phenological analyses did not show significant site differences that were related to latitude for any of five phenophases, but indicated that flowering may be nearly synchronous in Texas, Florida and St. Croix. Statistical analyses of one phenophase, the initial appearance of visible floral buds, and the temperatures antecedent to this phenophase indicated that flowering in St. Croix is preceded by a significantly higher temperature progression than that in Florida or in Texas. Under controlled conditions, plants of Texas were induced to flower at temperatures at or below 23°C and to reach anthesis at 23°C, but plants of more tropical origin, St. Croix and Mexico, flowered less frequently and at a slightly higher temperature, 24–26°C. Plants of diverse origin produced flowers under continuous light, suggesting that photoperiod does not play a significant role in flowering phenology. The combined studies in the natural seagrass beds and in the laboratory under controlled conditions indicate that temperature responses for St. Croix plants are probably genotypically different from those of Florida and Texas.